Development of a Single-Cell Technique to Increase Yield and Use of Gastrointestinal Cancer Organoids for Personalized Medicine Application.

Abstract

BACKGROUND:

Organoids are excellent 3-dimensional in vitro models of gastrointestinal cancers. However, patient-derived organoids (PDOs) remain inconsistent and unreliable for rapid actionable drug sensitivity testing due to size variation and limited material.

STUDY DESIGN:

On day10/passage 2 after standard creation of organoids, half of PDOs were dissociated into single-cells with TrypLE Express Enzyme/DNase I and mechanical dissociation; and half of PDOs were expanded by the standard technique. Hematoxylin and eosin and immunohistochemistry with CK7 and CK20 were performed for characterization. Drug sensitivity testing was completed for single-cells and paired standard PDOs to assess reproducibility.

RESULTS:

After 2 to 3 days, >50% of single-cells reformed uniform miniature PDOs (∼50 μm). We developed 10 PDO single-cell lines (n = 4, gastric cancer, [GC]; and n = 6, pancreatic ductal adenocarcinoma, [PDAC]), which formed epithelialized cystic structures and by IHC, exhibited CK7(high)/CK20(low) expression patterns mirroring parent tissues. Compared with paired standard PDOs, single-cells (n = 2, PDAC; = 2, GC) showed similar architecture, albeit smaller and more uniform. Importantly, single cells demonstrated similar sensitivity to cytotoxic drugs to matched PDOs.

CONCLUSIONS:

PDO single-cells are accurate for rapid clinical drug testing in gastrointestinal cancers. Using early passage PDO single-cells facilitates high-volume drug testing, decreasing time from tumor sampling to actionable clinical decisions, and provides a personalized medicine platform to optimally select drugs for gastrointestinal cancer patients.

• Authors: Mei Gao, Megan M Harper, Miranda Lin, Shadi A Qasem, Reema A Patel, Samuel H Mardini, Moamen M Gabr, Michael J Cavnar, Prakash K Pandalai, Joseph Kim
• Journal: Journal of the American College of Surgeons (J Am Coll Surg) Vol. 232 Issue 4 Pg. 504-514 (04 2021) ISSN: 1879-1190 [Electronic] United States
• PMID: 33253861 (Publication Type: Journal Article, Research Support, N.I.H., Extramural)
• Copyright: Copyright © 2020 American College of Surgeons. Published by Elsevier Inc. All rights reserved.

Chemical References

• Antineoplastic Agents

Topics

• Antineoplastic Agents (pharmacology, therapeutic use)
• Biopsy
• Cell Survival
• Drug Resistance, Neoplasm (genetics)
• Drug Screening Assays, Antitumor (methods)
• Gastrointestinal Neoplasms (drug therapy, genetics, pathology)
• Humans
• Organoids (drug effects, pathology)
• Precision Medicine (methods)
• Primary Cell Culture (methods)
• Reproducibility of Results
• Single-Cell Analysis (methods)
• Time Factors